Proviral latency remains an important barrier to eradication of HIV from infected individuals. Epigenetic processes are critical in the regulation of HIV gene expression. We hypothesize that arginine and lysine methylation plays an important silencing role in proviral latency and that small molecule inhibitors of methyltransferases can contribute therapeutically to the induction/eradication (l/E) of HIV in latently infected T cells. This hypothesis is supported by data in the literature and our own preliminary unpublished studies, in which we identified several methyltransferases (MTs) with suppressive activity against HIV transcription in latently infected T cells. The goal of Project 1.5 is to comprehensively study the role of MTs in persistent HIV infection and in close collaboration with investigators across the Martin Deianey Collaboratory to validate MTs as potential drug targets in the l/E of HIV proviral latency. Specific Aim 1: Comprehensively analyze small hairpin RNAs directed against lysine and arginine methyltransferases in J-Lat cells and primary cell culture models of HIV latency. We will perform a highly targeted shRNA screen of 11 known human arginine MTs, 24 known human lysine MTs and 38 new proteins identified based on domain homology with a conserved SET domain characteristic for lysine MTs to identify a set of MTs that enforce HIV latency as potential drug targets for l/E therapy. Specific Aim 2: Study the molecular mechanisms how methyltransferases control HIV latency. We will study the transcriptional activities of four suppressive MTs (PRMT1, PRMT6, Set1 and SETDB1) identified by our preliminary work. We will study the in wVo recruitment of these factors to the latent HIV LTR and identify substrates of these enzymes in latently infected T cells (histones. Tat, P-TEFb or NF-KB). Specific Aim 3: Test the effect of methyltransferase inhibitors on the reactivation of HIV transcription in latently infected T cells. We will evaluate the effect of known small molecule inhibitors of arginine and lysine MTs on HIV proviral latency in J-Lat cells and primary T cell models of latency. We will characterize the enzymes primarily targeted by these inhibitors and use this information for the development of effective and specific l/E of HIV, Collectively, these studies will provide novel molecular insight into the basic mechanisms governing HIV proviral latency and support the collective goal of this Collaboratory to produce compounds to be tested in the l/E of persistent HIV infection in animals and patients.